PXD008624

PXD008624 is an original dataset announced via ProteomeXchange.

Title	E2~dID: A versatile approach to identify substrates modified with ubiquitin and ubiquitin-like molecules by specific E3 enzymes
Description	Covalent modifications of proteins with ubiquitin and ubiquitin-like molecules are instrumental to most, if not all biological processes. However, identifying the E3 ligase responsible for these modifications remains a major bottleneck in ubiquitin research. Here, we have developed an E2-thioester-driven identification (E2~dID) method for the targeted identification of substrates of specific E2 and E3 enzyme pairs. E2~dID exploits the central position of E2 conjugating enzymes in the ubiquitination cascade and provides in vitro generated biotinylated E2~ubiquitin thioester conjugates as the sole source for ubiquitination in extracto. This enables purification and identification of modified proteins by mass spectrometry under stringent conditions independently of the biological source of the extract. We demonstrate the sensitivity and specificity of E2-dID by identifying and validating substrates of the APC/C in human cells. Finally, performing E2~dID with SUMO in S. cerevisiae we show that E2-dID can be easily adapted to other ubiquitin-like modifiers and experimental models.
HostingRepository	PRIDE
AnnounceDate	2019-04-03
AnnouncementXML	Submission_2019-04-03_03:11:02.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	James Wright
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606; scientific name: Saccharomyces cerevisiae (Baker's yeast); NCBI TaxID: 4932;
ModificationList	TMT6plex-126 reporter+balance reagent acylated residue; ubiquitination signature dipeptidyl lysine; sumoylated lysine; monohydroxylated residue; acetylated residue; iodoacetamide derivatized residue; deamidated residue
Instrument	Orbitrap Fusion Lumos

Revision	Datetime	Status	ChangeLog Entry
0	2018-01-09 05:24:02	ID requested
⏵ 1	2019-04-03 03:11:03	announced

Bakos G, Yu L, Gak IA, Roumeliotis TI, Liakopoulos D, Choudhary JS, Mansfeld J, An E2-ubiquitin thioester-driven approach to identify substrates modified with ubiquitin and ubiquitin-like molecules. Nat Commun, 9(1):4776(2018) [pubmed]

curator keyword: Biological

submitter keyword: Ubiquitin, AP-MS, TMT

Jyoti Choudhary
contact affiliation	The Institute of Cancer Research
contact email	jyoti.choudhary@icr.ac.uk
lab head
James Wright
contact affiliation	Functional Proteomics, Institute Cancer Research & Proteomic Mass Spectrometry, Wellcome Trust Sanger Institute
contact email	jw13@sanger.ac.uk
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2019/04/PXD008624

PRIDE project URI

• PRIDE
  1. PXD008624
     1. Label: PRIDE project
     2. Name: E2~dID: A versatile approach to identify substrates modified with ubiquitin and ubiquitin-like molecules by specific E3 enzymes